GSM6619631: Mouse 2 scRNA; Mus musculus; RNA-Seq
The two tumors were isolated and minced with scalpels on ice. For enzymatic dissociation, 5 ml Papain solution (in DMEM/F12) supplmented with 1000 U DNase ( was added to each sample, followed by incubation at 37°C in 5% CO2 for 30 min. The cell solution was then transferred to a 40 µm cell strainer. Red blood cells were lysed with ACK lysis buffer (5 min. at 4°C). The resulting single cell suspensions were stained with 7-aminoactin mycin D (7-AAD). Non-viable, 7-AAD-positive cells were removed by fluorescence-activated cell sorting before viable single cells were further processed using 10x Genomics Chromium Next GEM Single Cell 3' Technology. Approximately 10,000 single cells of each sample were processed for scRNA-seq using the Chromium Next GEM Single Cell 3' technology (10X Genomics) according to the manufacturer's instructions. In brief, single-cell GEMs (gel beads in emulsion) were generated by the Chromium Controller, followed by GEM reverse transcription, a cleanup using Dynabeads, cDNA amplification, cleanup and size selection using SPRIselect beads, and construction of the sequencing library using Library & Gel Bead Kit v3.1, the Chromium Next GEM Chip G and the Single Index Kit T. Quality, purity, average size and concentration of cDNA and libraries were determined by measurements on a Tapestation 2200 (Agilent Technologies). Indexed libraries were sequenced using an Illumina NextSeq 2000 sequencing platform.